angiogenin and Hypoxia

Angiogenesis is essential for tumor growth and metastasis. Many signaling pathways are involved in regulating tumor angiogenesis, with the vascular endothelial growth factor pathway being of particular interest. The recognition of the heterogeneity in tumor vasculature has led to better predictions of prognosis through differential analyses of the vasculature. However, the clinical benefits from antiangiogenic therapy are limited, because many antiangiogenic agents cannot provide long-term survival benefits, suggesting the development of drug resistance. Activation of the hypoxia and c-Met pathways, as well as other proangiogenic factors, has been shown to be responsible for such resistance. Vessel co-option could be another important mechanism. For future development, research to improve the efficacy of antiangiogenic therapy includes (a) using tumor-derived endothelial cells for drug screening; (b) developing the drugs focusing on specific tumor types; (c) developing a better preclinical model for drug study; (d) developing more accurate biomarkers for patient selection; (e) targeting the c-Met pathway or other pathways; and (f) optimizing the dose and schedule of antiangiogenic therapy. In summary, the future of antiangiogenic therapy for cancer patients depends on our efforts to develop the right drugs, select the right patients, and optimize the treatment conditions.

Topics: Angiogenesis Inhibitors; Cell Proliferation; Humans; Hypoxia; Hypoxia-Inducible Factor 1, alpha Subunit; Microvessels; Neoplasm Metastasis; Neoplasms; Neovascularization, Pathologic; Proto-Oncogene Proteins c-met; Ribonuclease, Pancreatic; Signal Transduction; TOR Serine-Threonine Kinases; Vascular Endothelial Growth Factors

Retinal neovascularization (RNV) is a type of serious vision‑threating disease, commonly induced by hypoxia of ischemic retinopathy, which happens in various ocular diseases including diabetic retinopathy and retinopathy of prematurity. In clinical work, anti‑VEGF therapy is the preferred strategy for treating RNV. However, not all cases are sensitive to anti‑VEGF injection. It is urgent and necessary to develop novel targets for inhibiting neovascularization in ocular diseases. Angiogenin (ANG) and brain‑derived neurotrophic factor (BDNF) are implicated in angiogenesis, although their regulation and effects in RNV remain to be elucidated. microRNA (miRNA) is a type of small non‑coding RNA, which can modulate targets by degrading transcripts or inhibiting protein translation. In the present study, miRNA‑mediated modulation of ANG and BDNF was explored in an oxygen‑induced retinopathy mouse model and human retinal microvascular endothelial cells (HRECs) under hypoxia. The results showed that downregulation of miR‑182‑5p and upregulation of ANG and BDNF were found

Topics: Animals; Brain-Derived Neurotrophic Factor; Disease Models, Animal; Endothelial Cells; Female; Gene Expression Regulation; Humans; Hypoxia; Mice; Mice, Inbred C57BL; MicroRNAs; Neovascularization, Pathologic; Oxygen; Pregnancy; Retinal Neovascularization; Retinal Vessels; Ribonuclease, Pancreatic

A major mediator of angiogenesis is angiogenin, which is expressed in the early phase of healing in oral tissue engineering strategies. It is unclear how angiogenin is regulated in the periodontal tissue. The objective of this study was to reveal the regulation of angiogenin in response to hypoxia and the hypoxia mimetic agent l-mimosine in periodontal fibroblasts.. Human fibroblasts of the periodontal ligament (PDLF) and the gingiva (GF) in monolayer and spheroid cultures were exposed to hypoxia or l-mimosine. The production of angiogenin was evaluated at mRNA and protein levels with reverse transcription quantitative polymerase chain reaction and enzyme-linked immunosorbent assays, respectively. Echinomycin, an inhibitor of hypoxia-inducible factor (HIF)-1 activity, was used to test the involvement of HIF-1.. Our data show that hypoxia and l-mimosine can increase angiogenin mRNA and protein levels in PDLF monolayer cultures. In GF monolayer cultures, we found an increase of angiogenin at the mRNA level in response to hypoxia. The increase of angiogenin can be blocked by inhibition of HIF-1 signaling via echinomycin. In PDLF and GF spheroid cultures, the impact of hypoxia and l-mimosine did not reach the level of significance.. Hypoxia and the hypoxia mimetic agent l-mimosine can increase the production of angiogenin via HIF-1 signaling in PDLF monolayer cultures but not in spheroid cultures. GF were less sensitive to the impact of hypoxia and l-mimosine. Overall, these results suggest a link between hypoxia, HIF-1 signaling and angiogenin in the periodontium.

Topics: Cells, Cultured; Fibroblasts; Humans; Hypoxia; Hypoxia-Inducible Factor 1, alpha Subunit; Mimosine; Periodontal Ligament; Ribonuclease, Pancreatic

The disruption of appropriate cellular stress responses is implicated in the pathogenesis of different neurological disorders including ischemic injury. Early diagnosis and treatment are often associated with better prognosis in ischemic stroke patients. Thus, there is an urgent need to improve the speed and accuracy of stroke diagnosis by developing highly sensitive stroke biomarkers. We recently reported that transfer RNA (tRNA) was involved in cell stress response pathways. Under cell stress conditions, mature tRNA is cleaved by a specific ribonuclease, angiogenin, generating tRNA-derived stress-induced RNA (tiRNA). To study tiRNA generation in an in vitro model of ischemic-reperfusion injury, we used the rat neuronal cell line, PC12, in combination with analysis of SYBR staining and immuno-northern blotting using anti-1-methyladenosine antibody, which detects 1-methyladenosine (m

Topics: Adenosine; Animals; Biomarkers; Cell Survival; Glucose; Hypoxia; Ischemia; Neurons; Oxygen; PC12 Cells; Rats; Reperfusion Injury; Ribonuclease, Pancreatic; RNA, Transfer; Stress, Physiological; Stroke

Hypoxia induces complex cellular responses that are mediated by a key transcription factor, hypoxia-inducible factor-1 (HIF-1). HIF-1 promotes production of cytokines and angiogenic factors and contributes to recovery of injured tissues. In the present study, expressions of angiogenin (ANG) and vascular endothelial growth factor (VEGF), which are potent angiogenic factors in mammalian tissues, were examined in immortalized fibroblasts exposed to hypoxia. After 24 h of exposure to hypoxia, ANG and VEGF mRNAs expressions were significantly elevated in periodontal ligament (PDL) fibroblasts but not in embryonic fibroblasts. Hypoxia also increased productions of ANG and VEGF proteins in PDL fibroblasts. HIF-1α mRNA expression was not affected by hypoxia in either fibroblast, although HIF-1α protein expression was enhanced after exposure to hypoxia. Treatment of PDL fibroblasts with dimethyloxaloylglycine, a prolyl hydroxylase inhibitor that stabilizes the HIF-1α protein, significantly increased expressions of ANG and VEGF mRNAs under normoxia. This suggests that stabilization of HIF-1α is crucial for upregulation of ANG and VEGF in PDL fibroblasts. These results indicate that, under hypoxic conditions, HIF-1α upregulates synthesis of ANG and VEGF in PDL fibroblasts and promotes angiogenesis.

Topics: Blotting, Western; Cell Line; Cells, Cultured; Cytokines; Fibroblasts; Fluorescent Antibody Technique; Humans; Hypoxia; Hypoxia-Inducible Factor 1; Periodontal Ligament; Real-Time Polymerase Chain Reaction; Ribonuclease, Pancreatic; RNA, Messenger; Up-Regulation; Vascular Endothelial Growth Factor A

Angiogenin is a key molecule in the healing process which has been successfully applied in the field of regenerative medicine. The role of angiogenin in dental pulp regeneration is unclear. Here we aimed to reveal the impact of the hypoxia mimetic agent L-mimosine (L-MIM) and hypoxia on angiogenin in the dental pulp.. Human dental pulp-derived cells (DPC) were cultured in monolayer and spheroid cultures and treated with L-MIM or hypoxia. In addition, tooth slice organ cultures were applied to mimic the pulp-dentin complex. We measured angiogenin mRNA and protein levels using qPCR and ELISA, respectively. Inhibitor studies with echinomycin were performed to reveal the role of hypoxia-inducible factor (HIF)-1 signaling.. Both, L-MIM and hypoxia increased the production of angiogenin at the protein level in monolayer cultures of DPC, while the increase at the mRNA level did not reach the level of significance. The increase of angiogenin in response to treatment with L-MIM or hypoxia was reduced by echinomycin. In spheroid cultures, L-MIM increased angiogenin at protein levels while the effect of hypoxia was not significant. Angiogenin was also expressed and released in tooth slice organ cultures under normoxic and hypoxic conditions and in the presence of L-MIM.. L-MIM and hypoxia modulate production of angiogenin via HIF-1 differentially and the response depends on the culture model. Given the role of angiogenin in regeneration the here presented results are of high relevance for pre-conditioning approaches for cell therapy and tissue engineering in the field of regenerative endodontics.

Topics: Cells, Cultured; Dental Pulp; Echinomycin; Enzyme-Linked Immunosorbent Assay; Humans; Hypoxia; Mimosine; Reverse Transcriptase Polymerase Chain Reaction; Ribonuclease, Pancreatic; RNA, Messenger

Microvesicles (MVs) derived from mesenchymal stem cells (MSCs) have been shown to promote angiogenesis. This study was aimed to shed a light on the mechanisms by analyzing the angiogenesis-promoting compositions of MSC-MVs. Also we try to figure out the impact of hypoxia on angiogenesis.. MVs were isolated from the culture supernatants of MSCs under hypoxia/normoxia and serum-deprivation condition. The morphological features of MVs were revealed by an electron microscope and the origin of the MVs was identified by a bead-bound assay. An antibody array was used to analyze the expression of angiogenic cytokines from MVs and the parent MSCs as well. The major candidate factors were screened and the results were validated by immune blotting.. MSC-MVs were around 80 nm in diameter. They expressed CD29, CD44, and CD73, but not CD31 and CD45. Antibody array showed that both MSCs and MVs expressed many angiogenesis-promoting biomolecules, including interleukin-6 (IL-6), basic fibroblast growth factors (bFGF), and recptor of urokinase-type plasminogen activator (UPAR). MSC-MVs contained angiogenin, vascular endothelial growth factor (VEGF), monocyte chemotactic protein-1 (MCP-1) and the receptor-2 for vascular endothelial growth factor at higher levels than the parent MSCs. Under hypoxic condition most cytokines were expressed in greater quantity than normoxic in MSCs while in MVs there was no significant difference between hypoxic and normoxic conditions except UPAR, Angiogenin, VEGF, IGF, Tie-2/TEK, and IL-6 which were higher in MVs under hypoxic conditions than those in normoxic condition.. Upon serum-deprivation condition, MSCs could secrete MVs that contain a variety of factors contributing to their angiogenesis-promoting function. And among them, Angiogenin, VEGF, MCP-1, VEGF R2 might be of greater importance than the other cytokines. Also UPAR, Angiogenin, VEGF, IGF, Tie-2/TEK, IL-6 might be responsible for hypoxia-augmented proangiogenic effects of MVs.

Topics: Analysis of Variance; Blotting, Western; Chemokine CCL2; Cytokines; Humans; Hypoxia; Mesenchymal Stem Cells; Microscopy, Electron, Scanning; Microvessels; Neovascularization, Physiologic; Ribonuclease, Pancreatic; Umbilical Cord; Vascular Endothelial Growth Factor A

Angiogenin (ANG) is a potent angiogenic factor that is up-regulated by hypoxia. ANG expression is well documented in normal tissues and in common tumours, but its expression has not been reported in the normal human kidney or in Wilms' tumours (WT). We examined ANG expression in WTs, human fetal kidney (FK) and childhood kidney (NK) samples and studied its relationship with microvascular density (MVD) and with three other hypoxia-induced angiogenic factors: lactate dehydrogenase A (LDHA), vascular endothelial growth factor (VEGFA) and BHLHE40 (basic helix-loop-helix transcription factor E40). Total ANG protein levels were significantly lower in WTs when compared with those in 15 matched-paired NKs. ANG immunoreactivity was observed in the glomeruli, proximal tubules and vessels in the FKs and NKs, indicating that ANG plays a physiological role in the human kidney. ANG cellular localization and distribution in 27 WTs reflected the pattern observed in the FKs. ANG colocalized with LDHA in the perinecrotic areas of untreated WTs suggesting up-regulation by hypoxia. There was a significant correlation between CD31-MVD and ANG-MVD. ANG, CD31, VEGFA and BHLHE40 mRNA levels were significantly lower in 15 WTs compared with matched-paired NKs. Univariable and multivariable statistical analyses showed significant correlations between ANG and CD31, ANG and BHLHE40 mRNAs and a weaker relationship between ANG and VEGFA mRNAs. ANG expression in WTs recapitulates that seen during nephrogenesis, and correlation with CD31-MVDs and mRNAs is consistent with a contribution to angiogenesis in WTs. Our study contributes to the understanding of angiogenesis during development and in WTs.

Topics: Angiogenesis Inducing Agents; Basic Helix-Loop-Helix Transcription Factors; Child; Cluster Analysis; Female; Fetus; Gene Expression Regulation; Homeodomain Proteins; Humans; Hypoxia; Isoenzymes; Kidney; L-Lactate Dehydrogenase; Lactate Dehydrogenase 5; Platelet Endothelial Cell Adhesion Molecule-1; Pregnancy; Ribonuclease, Pancreatic; RNA, Messenger; Up-Regulation; Vascular Endothelial Growth Factor A; Wilms Tumor

Our objective was to assess the regulation of the hypoxia response of angiogenic and inflammatory factors from 76 cerebrospinal fluids (CSF) of sporadic amyotrophic lateral sclerosis (ALS) patients with different respiratory status. We first analysed the hypoxia response capacity by measuring CSF levels of angiogenin (ANG), VEGF, angiopoietin-2 (ANG-2) and PGE-2 in 40 ALS patients according to their hypoxaemia level and compared it with 40 neurological controls. We then compared the ANG, VEGF, EPO and ANG-2 CSF levels of 36 other ALS patients, divided into three groups with either 1) normoxaemia, 2) intermittent desaturation in the absence of hypoxaemia, or 3) chronic hypoxaemia with or without desaturation. We demonstrated a lack of up-regulation of both ANG and VEGF during hypoxaemia in ALS, compared with hypoxaemic controls. In contrast, PGE-2 and ANG-2 levels were increased in both hypoxaemic ALS patients and controls. ANG and VEGF levels did not increase in patients with long disease durations and with intermittent or chronic hypoxaemia. ANG-2 and EPO levels were up-regulated early in intermittent hypoxaemia and late in chronic hypoxaemia, respectively. Our results suggest alteration of the HIF-1alpha-mediated response to hypoxia during sporadic ALS, whereas the NFK-B pathway seems early activated.

Topics: Aged; Amyotrophic Lateral Sclerosis; Angiopoietin-2; Dinoprostone; Erythropoietin; Humans; Hypoxia; Inflammation; Middle Aged; Oxygen; Respiration; Ribonuclease, Pancreatic; Vascular Endothelial Growth Factor A

Amyotrophic lateral sclerosis (ALS) is a fatal progressive neurodegenerative disease characterized by the selective death of motor neurons in the motor cortex, brain stem and spinal cord. Recently, missense variants in the angiogenin gene (ANG), an angiogenic factor expressed in ventral horn motor neurons that is up-regulated by hypoxia, have been found in ALS patients of Irish/Scottish, North American, Italian, French and Dutch descent. To investigate the role of ANG in the German population, we screened for mutations by sequencing the entire coding region of the ANG gene in a large sample of 581 German ALS cases and 616 sex- and age-matched healthy controls. We identified two heterozygous missense variants, F(-13)L and K54E, in two German sporadic ALS cases but not in controls. Both missense variants are novel and have not been previously found in ALS cases. Our results suggest that missense variants in the ANG gene play a role in ALS in the German population and provide further evidence to support the hypothesis that angiogenic factors up-regulated by hypoxia are involved in the pathophysiology of ALS.

Topics: Amyotrophic Lateral Sclerosis; Central Nervous System; Cohort Studies; DNA Mutational Analysis; Female; Genetic Markers; Genetic Predisposition to Disease; Genetic Testing; Genetic Variation; Genotype; Germany; Heterozygote; Humans; Hypoxia; Male; Middle Aged; Mutation, Missense; Oxidative Stress; Ribonuclease, Pancreatic; White People

Genetic variations in two hypoxia-inducible angiogenic genes, VEGF and ANG, have been linked with sporadic amyotrophic lateral sclerosis (SALS). Common variations in these genes may reduce the levels or functioning of their products. VEGF and ANG belong to a larger group of angiogenic genes that are up-regulated under hypoxic conditions. We hypothesized that common genetic variation across other members of this group may also predispose to sporadic ALS. To screen other hypoxia-inducible angiogenic genes for association with SALS, we selected 112 tagging single nucleotide polymorphisms (tgSNPs) that captured the common genetic variation across 16 VEGF-like and eight ANG-like hypoxia-inducible genes. Screening for association was performed in 270 Irish individuals with typical SALS and 272 ethnically matched unrelated controls. SNPs showing association in the Irish phase were genotyped in a replication sample of 281 Swedish sporadic ALS patients and 286 Swedish controls. Seven markers showed association in the Irish. The one modest replication signal observed in the Swedish replication sample, at rs3801158 in the gene inhibin beta A, was for the opposite allele vs. the Irish cohort. We failed to detect association of common variation across 24 candidate hypoxia-inducible angiogenic genes with SALS.

Topics: Aged; Amyotrophic Lateral Sclerosis; Female; Genetic Predisposition to Disease; Genetic Testing; Genotype; Humans; Hypoxia; Ireland; Male; Middle Aged; Polymorphism, Single Nucleotide; Ribonuclease, Pancreatic; Sequence Analysis, DNA; Sweden; Vascular Endothelial Growth Factor A

The role of hypoxia responsive genes in the pathogenesis of ALS was first suggested when deletions of the hypoxia-responsive element of vascular endothelial growth factor (VEGF) promoter caused a motor neuron disease phenotype in mice. The discovery of ALS-associated mutations in ANG, a hypoxia responsive gene coding for the protein angiogenin, has further supported this pathogenic mechanism in human ALS. In endothelium, angiogenin can regulate expression of VEGF. To date, the patterns of serum angiogenin expression among patients with ALS have not been assessed.. Serum angiogenin and VEGF levels were quantified at diagnosis in 79 patients with definite or probable ALS and 72 healthy controls, using a quantitative sandwich enzyme-linked immunoassay.. Patients with ALS exhibited higher serum angiogenin (p = 0.006) but not VEGF (p = 0.55) levels than matched control subjects. Subgroup analysis showed a greater elevation in angiogenin levels for spinal- (p < 0.001) than bulbar- (p = 0.11) onset ALS vs controls. At 12 months, angiogenin levels remained elevated. No correlation was noted between angiogenin and VEGF levels (r = -0.08, p = 0.49) in ALS patient serum.. These data suggest a modest elevation in serum angiogenin in ALS at diagnosis. Further investigation will be required to assess the utility of serum angiogenin as a biomarker for ALS and as a predictor of disease progression.

Topics: Amyotrophic Lateral Sclerosis; Biomarkers; Female; Humans; Hypoxia; Male; Microcirculation; Neovascularization, Physiologic; Predictive Value of Tests; Ribonuclease, Pancreatic; Spinal Cord; Survival Rate; Up-Regulation; Vascular Endothelial Growth Factor A

Many processes that are involved in cellular invasion, including blastocyst implantation, placental development, and rapidly growing tumors, occur in reduced oxygen environments. It has been surmised that oxygen tension could regulate the cytotrophoblast ability to differentiate and, as a consequence, to express proteins that are critical for placentation. The objective of the current investigation was therefore to test the hypothesis that placental tissues and trophoblast cells in culture, under low oxygen tension, release angiogenic factors that could affect vascular behavior and invasive potential, thus providing a link between abnormal placentation and maternal vascular abnormality.. Functionally active term placental explant culture and trophoblast cultures were used to demonstrate the secretion profiles of angiogenin and vascular cell adhesion molecule-1 (VCAM-1), and the real-time quantitative reverse transcriptase polymerase chain reaction (RT-PCR) technique was employed to demonstrate the mRNA expression under both normoxic and hypoxic conditions.. A significant increase in the secretion (P <.01) and mRNA expression (P <.01) of angiogenin and a significant decrease in the secretion (P <.04) and mRNA expression (P <.03) of VCAM-1 from both term placental explants and trophoblast cultures subjected to hypoxia in vitro were observed.. Because the primary defect in uteroplacental insufficiency is placental maldevelopment probably associated with hypoxia in situ, this study provides molecular evidence to indicate that the differential expression and secretion of angiogenic factors may play an important role in these pathologic conditions.

Topics: Cell Culture Techniques; Female; Gene Expression Profiling; Humans; Hypoxia; Neovascularization, Physiologic; Oxygen; Placenta; Placentation; Pregnancy; Pregnancy Complications; Reverse Transcriptase Polymerase Chain Reaction; Ribonuclease, Pancreatic; RNA, Messenger; Trophoblasts; Up-Regulation; Vascular Cell Adhesion Molecule-1

Knowledge of the rapidly developing hierarchy of controls affecting vascular development in placenta is required to understand how the growth factors and their receptor-mediated signals actually produce vessels. At the cell biological level, these events clearly require stable interactions between the cells, and cells with the surrounding ECM. The objective of the study was to understand the role of integrins and ECM on the expression and secretion of angiogenin in placentas and from trophoblasts in culture. Functionally active term placental explant culture and trophoblast cultures were used to demonstrate the differential secretion profile of angiogenin and real-time quantitative RT-PCR to demonstrate the mRNA expression in the presence or absence of ECM proteins. In this study, a significant increase in expression and secretion of angiogenin occurred in the presence of vitronectin (VN) and fibronectin (FN). Using antibody-blocking experiments it was also demonstrated that the angiogenin secretion is mediated by placental integrins, alpha(V)beta3 and alpha5beta1. In addition, exposure to hypoxic conditions resulted in diminished angiogenin secretion in the presence of both ECMs suggesting that angiogenin expression in the presence of ECM is modulated by local O2 concentration. In conclusion, this study provides evidence for the regulatory role of ECM and integrins on the mRNA expression and secretion of angiogenin in human placenta. ECMs may have a pivotal role in enhancing secretion of this peptide necessary for placental angiogenesis and provides the impetus as additional targets for the control of angiogenesis in pathological pregnancy.

Topics: Cell Line, Tumor; Cells, Cultured; Extracellular Matrix; Female; Gene Expression Regulation; Humans; Hypoxia; Oxygen; Placenta; Pregnancy; Ribonuclease, Pancreatic; RNA, Messenger; Tissue Culture Techniques; Trophoblasts

DEC1, also known as SHARP-2 or Stra13, plays important roles in embryonic development, proliferation, apoptosis and cell differentiation in the mouse. DEC1 was recently identified as hypoxically induced in cDNA microarray studies of the human renal carcinoma cell line RCC4, to be regulated through hypoxia-inducible factor (HIF)-1alpha and via HIF-1alpha, able to block adipocyte differentiation. Nevertheless, its distribution and role in hypoxia and differentiation in human breast cancer are unknown. We therefore examined the pattern and level of expression of DEC1 using immunohistochemistry in whole tissue sections in normal, in situ and invasive breast carcinomas, and correlated the level of expression of DEC1 and clinicopathological factors and hypoxic tumour markers in 253 invasive carcinomas on tissue microarrays. We observed an increase in DEC1 expression during progression from normal to in situ and invasive carcinoma. Expression was not restricted to the tumour cell element but was also observed in endothelial, fibroblasts and inflammatory cells. There was a significant positive correlation between DEC1 and tumour grade (P=0.01), HIF-1alpha (P=0.04) and the hypoxically regulated gene angiogenin (P<0.0001), but no significant associations were observed with patient age (P=0.15), lymph node status (P=0.8), tumour size (P=0.3), oestrogen receptor (P=0.45), epidermal growth factor receptor (P=0.27) or Chalkley vessel count (P=0.45). There was no difference in relapse-free (P=0.84) or overall (P=0.78) survival. These findings suggest that DEC1 plays an important role in the progression to invasive breast cancer and that it may provide a mechanism by which hypoxia blocks tumour differentiation, and may contribute to a more aggressive phenotype. Reversing this phenotype may alter the biological behaviour of individual tumours.

Topics: Adult; Basic Helix-Loop-Helix Transcription Factors; Biomarkers, Tumor; Breast; Breast Neoplasms; Carcinoma in Situ; Cell Transformation, Neoplastic; Disease Progression; Female; Homeodomain Proteins; Humans; Hypoxia; Hypoxia-Inducible Factor 1, alpha Subunit; Immunohistochemistry; Middle Aged; Prognosis; Ribonuclease, Pancreatic; Transcription Factors; Tumor Suppressor Proteins

Angiogenesis is thought to be crucial for normal physiology of the endometrium, where dynamic vascular remodeling occurs during the menstrual cycle and pregnancy. We investigated the presence of angiogenin, a potent inducer of angiogenesis, and the regulatory mechanisms of its production in the human endometrium. Western blot analysis demonstrated that angiogenin protein expression increased by 3- to 4-fold in the endometrium in the mid and late secretory phases and in early gestation relative to that during the proliferative phase. Quantitative mRNA analysis showed the similar tendency in the expression of angiogenin mRNA in the endometrium, with the highest levels observed in the mid and late secretory phases and early gestation. An immunohistochemical study showed that angiogenin was expressed in both stromal cells and epithelial cells, with indistinguishable intensity between these cells regardless of phases of the menstrual cycle. In support of the Western blot analysis, the intensity of staining appeared to be highest in the mid to late secretory phases relative to other phases. Consistent with these in vivo results, decidualized cultured stromal cells, after treatment with progesterone or progesterone plus E2, exhibited the capacity to secrete significantly increased amounts of angiogenin compared with untreated or E2 alone-treated control group. Both the treatment with (Bu)2cAMP and hypoxic conditions stimulated angiogenin secretion by stromal cells. For isolated epithelial cells, hypoxia stimulated angiogenin secretion, whereas (Bu)2cAMP had no appreciable effect. In summary, we demonstrated the presence of angiogenin in human endometrium and its possible local regulatory factors, such as progesterone, cAMP, and hypoxia. These findings along with its enhanced expression in the endometrium in the secretory phase and in decidual tissues raise the possibility that angiogenin may play a role in establishing pregnancy.

Topics: Blotting, Western; Cyclic AMP; Decidua; Endometrium; Epithelial Cells; Female; Humans; Hypoxia; Immunohistochemistry; In Vitro Techniques; Menstrual Cycle; Reverse Transcriptase Polymerase Chain Reaction; Ribonuclease, Pancreatic; RNA, Messenger; Stromal Cells

Angiogenesis is an essential event during the development of the ovarian follicle and ensuing formation of the corpus luteum. We investigated the presence of angiogenin, a potent inducer of angiogenesis, and the regulatory mechanisms of its production in the human ovary. Follicular fluid (FF) and granulosa cells (GCs) were collected from women undergoing in vitro fertilization and embryo transfer. The presence of angiogenin in FF and GCs was demonstrated by Western blot analysis. The production of angiogenin by cultured GCs was stimulated with the addition of human CG or cAMP or under the hypoxic milieu. Concentrations of angiogenin in FF from an individual follicle were positively correlated with those of progesterone, but not estradiol and testosterone. Given the presence of angiogenin in FF and up-regulation of its production by human CG and hypoxia, it seems logical to assume that angiogenin may play a role as a local angiogenic factor in the human ovary.

Topics: Adult; Blotting, Western; Bucladesine; Cells, Cultured; Chorionic Gonadotropin; Cyclic AMP; Female; Follicular Fluid; Granulosa Cells; Hormones; Humans; Hypoxia; Reverse Transcriptase Polymerase Chain Reaction; Ribonuclease, Pancreatic; Up-Regulation